Despite being the primary enzyme involved in methamphetamine (meth) metabolism, cytochrome P450-2D6 (CYP2D6) has not been studied (outside of our own work) with regard to its role in meth-related brain dysfunction. Our preliminary data with a small clinical sample suggest that genetic differences coding for the activity of CYP2D6 are associated with differential vulnerability to neurocognitive problems in meth users. Individuals with genotypes corresponding to diminished metabolism seem to incur less neuropsychological impairment than those with normal metabolism. This intriguing finding implicates the metabolic products of methamphetamine in brain injury. It also suggests a way to identify individuals are at increased risk of meth-related brain problems. Our group has described additive negative effects of meth and HIV on brain function. There is evidence that CYP2D6 activity may be altered in HIV, such that the genotype does not always correspond with the actual metabolic activity phenotype. Additionally, there are medications that are widely prescribed to HIV patients that are substrates and inhibitors of CYP2D6 and thus may affect meth metabolism if taken concurrently. These include serotonin reuptake inhibitors for depression as well as the antiretroviral ritonavir, which is used to boost a majority of cART regimens. There are reports in the literature of prolonged psychoactive drug effects in HIV patients taking ritonavir. To understand the mechanisms by which differences in CYP2D6 activity may contribute to meth-related brain dysfunction, we propose to generate a naturalistic in vitro model of neural injury associated with meth exposure in the context of HIV. We will use human mixed neuroglial cultures derived from fetal tissue donation. As such, the cultures can be genotyped. We propose to treat the cultures with meth and HIV, and (1) measure metabolite formation; (2) determine whether CYP2D6 activity / metabolite quantities are related to amount of cellular injury; and (3) examine how these results are affected by exposure of the cultures to clinically relevant CYP2D6 inhibitors (paroxetine and ritonavir). We expect that cultures with extensive (wildtype) metabolizer genotypes will generate the largest quantities of meth metabolites and will in turn show the greatest degree of cellular injury, followed by intermediate and then poor metabolizers. In preliminary work, we have demonstrated the feasibility of the various components of the proposed experiments. Findings stand to confirm our early human studies and provide a mechanism of meth-related injury in the context of HIV. Results could lead to the development of pharmacologic interventions to reduce brain injury as well as inform treatment choices for meth users with HIV.